The interaction of DNA with a novel cationic phospholipid transfection reag
ent, 1,2-dioleoyl-sn-glycero-3-ethylphosphocholine (EDOPC), was investigate
d by monitoring thermal effects, particle size, vesicle rupture, and lipid
mixing. By isothermal titration calorimetry, the heat of interaction betwee
n large unilamellar EDOPC vesicles and plasmid DNA was endothermic at both
physiological and low ionic strength, although the heat absorbed was slight
ly larger at the higher ionic strength. The energetic driving force for DNA
-EDOPC association is thus an increase in entropy, presumably due to releas
e of counterions and water. The estimated minimum entropy gain per released
counterion was 1.4 cal/mole-degrees K (about 0.7 kT), consistent with prev
ious theoretical predictions. All experimental approaches revealed signific
ant differences in the DNA-lipid particle, depending upon whether complexes
were formed by the addition of DNA to lipid or vice versa. When EDOPC vesi
cles were titrated with DNA at physiological ionic strength, particle size
increased, vesicles ruptured, and membrane lipids became mixed as the amoun
t of DNA was added up to a 1.6:1 (+:-) charge ratio. This charge ratio also
corresponded to the calorimetric end point. In contrast, when lipid was ad
ded to DNA, vesicles remained separate and intact until a charge ratio of 1
:1 (+:-) was exceeded. Under such conditions, the calorimetric end point wa
s 3:1 (+:-). Thus it is clear that fundamental differences in DNA-cationic
lipid complexes exist, depending upon their mode of formation. A model is p
roposed to explain the major differences between these two situations. Sign
ificant effects of ionic strength were observed; these are rationalized in
terms of the model. The implications of the analysis are that considerable
control can be exerted over the structure of the complex by exploiting vect
orial preparation methods and manipulating ionic strength.